The goal of the present research proposal is to attain a greater understanding of human platelet function by studying a critical protein component of platelets, the calcium pump. Since calcium has been shown to be a trigger for both platelet aggregation and the release of stored contents, the activity of the calcium pump is essential for maintaining normal platelet function. The project consists of two parts: (1) purification and characterization of the structure and function of the calcium pump protein, and (2) determination of the cellular localization of the pump. Preliminary work in this laboratory has shown that an antibody prepared against a calcium pump from rabbit skeletal muscle cross-reacts with a protein in solubilized human platelets. Since the antibody specifically precipitates the calcium pump activity, it appears that immunochemical procedures can be used to purify the calcium pump protein from human platelets. The purified pump can be characterized with respect to structure and enzymatic activity in detergent solution and in reconstituted phospholipid vesicles. The antibody can also be used to determine the subcellular localization of the pump protein. Determination of the properties and function of the Ca2+ pump in normal platelets will lead to a better understanding of platelet dysfunction. Platelet defects can lead to thrombosis, the final event in coronary and cerebral arterial occlusion. Thus, the present study will contribute to the general understanding of cardiovascular disease.